Prostate cancer (PCa) is the most common cancer in American men and the second highest cause of cancer related death in Western males with 189,000 new cases and over 30,000 deaths annually. HOXC6 and SOX4 are hormonally regulated developmental transcription factors that may confer a stem-cell like, androgen independent phenotype to prostate cancer cells. Our global gene expression profiling of prostate cancer patients has determined that HOXC6 and SOX4 are significantly upregulated in PCa and that their expression patterns are strongly correlated with the Gleason Score (or tumor grade). Immunohistochemistry shows that SOX4 protein is increased in prostate cancer cells, and immunoblots show increased HOXC6 protein levels in prostate cancer cells. Loss of HOXC6 and SOX4 expression induces apoptosis in prostate cancer cells, while overexpression induces increased proliferation and decreased apoptosis. Potential targets of HOXC6 and SOX4 may have roles in mitogenic signaling and cell cycle regulation. However, the biological effects of altered HOXC6 and SOX4 expression in prostate cells are not well understood. In this application, we propose to test the hypothesis that HOXC6 and/or SOX4 could be novel therapeutic targets for prostate cancer. Toward this goal, we will perturb HOXC6 and SOX4 expression in prostate cancer cells and measure apoptosis, proliferation rate, androgen-dependence, and growth in soft agar. We will also analyze the effects of in vivo siRNA treatment by co-injection of human prostate cancer cells and siRNAs into nude mice. We will further examine the effect of genetic loss of HOXC6 on prostate cancer incidence and progression using HOXC6-targeted null mice as a model system. In addition, candidate sets of HOXC6- and SOX4-regulated target genes that were identified in PCa patient tissue samples will be confirmed in prostate cells by expression profiling, quantitative real-time PCR, immunoblot, and chromatin immunoprecipitation (ChIP) assays. Finally, we will determine whether SOX4 levels are predictive of patient outcome using tissue arrays. Together these experiments will provide new insights into the mechanisms of prostate cancer progression and potentially validate HOXC6 and SOX4 as novel therapeutic targets for prostate cancer. [unreadable] [unreadable] [unreadable] [unreadable]